The present invention relates generally to novel antigens for developing antigen-binding molecules that are interactive with mammalian inhibins. More particularly, the present invention relates to immuno-interactive fragments of the (alpha)C portion of a mammalian inhibin alpha subunit and to variants and derivatives of these immuno-interactive fragments for producing novel antigen-binding molecules that recognize the said (alpha)C portion. The invention is also concerned with the use of these antigen-binding molecules for detecting a mammalian inhibin and for treating or preventing conditions associated with aberrant levels of a mammalian inhibin.
Inhibin is a dimeric glycoprotein produced by diverse tissues including the gonads, pituitary, brain, bone marrow, placenta, and adrenal gland. It was initially identified by its ability to inhibit the secretion of follicle stimulating hormone (FSH) by the pituitary (for reviews, see Vale et al., 1990, The inhibin/activin family of hormone and growth factors. In Peptide growth factors and their receptors: Handbook of Experimental Physiology 95:211-248 (Eds. Spom and Roberts) Springer-Verlag, Berlin; Burger, 1992, Reproductive Medicine Review 1:1-20; Baird & Smith, 1993, Oxford Rev. Reprod. Biol. 15:191-232). However, it was also found subsequently to be secreted by mucinous and granulosa cell cancers of the ovary. Thus, measurement of serum inhibin in women, particularly postmenopausal women, provides a good diagnostic test for detecting these cancers (Lapphorn et al., 1989, New England 0.1 Med. 321:790-793; Healy et al., 1993, N. Eng. J. Med. 329:1539-420) and for monitoring their recurrence after surgery. The mucinous and granulosa cell cancers represent 20-30% of all ovarian cancers. Serum inhibin is less effective as a marker of serous cancer, which is the major (40%) ovarian cancer. In contrast, a widely used cancer marker, CA125, is effective in the detection of serous cancers and less so with the mucinous and granulosa cell cancers.
Inhibin consists of two chains, the alpha subunit (made up of 3 regions, Pro, (alpha)N and (alpha)C) and either the (beta)A subunit (inhibin A) or (beta)B subunit (inhibin B), of varying molecular weight. Various inhibin assays with specificities directed towards different regions of the inhibin molecule have been developed for diagnosis of ovarian cancer.
Initial studies by Lapphorn et al. (1989, supra) and Healy et al. (1993, supra) suggested that measurement of serum inhibin by radioimmunoassay (RIA) which detects (alpha)C inhibin forms may be of diagnostic value in monitoring mucinous and granuloma cell tumours. Whilst this method is reliable, it is less sensitive and practical in comparison to two-site or sandwich antibody assays using, for example, colorimetric or fluorescent labels for detection.
A two-site immunofluorometric assay ((alpha)C IFMA) for the (alpha)C portion of the alpha subunit of inhibin has been developed by Robertson et al. (1996, J. Clin. Endocrinol. Metab. 81:669-676). This assay, which utilizes sheep polyclonal antisera and the fluorescent label Europium (Eu), detects all known inhibin alpha subunit-containing proteins. Compared to other inhibin assays specific for the alpha subunit or the (alpha)(beta) dimers (inhibin A and B), the (alpha)C IFMA and the (alpha)C RIA have been shown to be more effective in detecting different ovarian cancers (Robertson et al., 1999, Clin. Endocrinol. 50:381-387; ibid, Clin. Chemistry 45:651-658).
Robertson et al. (1999, Clin. Chemistry 45:651-658) have also shown that 89-90% of all ovarian cancers can be detected by the (alpha)C IFMA in combination with an immunoassay for the ovarian cancer marker CA125. This combined detection value was considerably higher than for each assay alone or a combination of CA125 with other inhibin assays, and is clinically useful in the diagnosis of the majority of ovarian cancers. Furthermore, in view of its increased sensitivity, the (alpha)C IFMA is able to detect the increase in serum inhibin associated with a recurrence of granulosa cell tumours at an earlier time following surgery. The earlier detection of the cancer is desirable for successful treatment.
Despite the clinical utility of the (alpha)C IFMA, the use of polyclonal antisera in this immunoassay or other types of multi-site assays in the diagnostic market is a disadvantage owing to the inherent limited supply of polyclonal antisera and the difficulties of quality control including specificity between antiserum batches. It would therefore be beneficial to utilize monoclonal antisera or other antigen-binding molecules where the stocks are potentially limitless and the quality can be more easily monitored.